Bacteriophage treatment of Campylobacter biofilms : formation of the carrier state life cycle

Campylobacter jejuni is a gram-negative thermotolerant microaerobic pathogen that causes human gastroenteritis worldwide. The production of extracellular polymeric substances to create a biofilm is a mechanism by which C. jejuni can protect itself from unfavourable environments, and is a contributory factor to the survival and transmission of the organism to farms animals and into the human food chain. Bacteriophages are natural predators of bacteria that have the potential for use as targeted biocontrol agents with the advantage that they can penetrate and affect bacteria embedded in biofilms. The effects of virulent Campylobacter-specific bacteriophages CP8 and CP30 on C. jejuni biofilms formed on glass by strains NCTC 11168, PT14 and HPC5 at 37 °C under microaerobic conditions were investigated. Independent bacteriophage treatment led to 1 to 3 Log10 CFU/cm2 reductions in the viable count 24 h postinfection compared with control levels. In contrast, bacteriophage applied under these conditions effected a reduction of less than 1 Log10 CFU/ml in planktonic cells. Resistance to bacteriophage in bacteria surviving bacteriphage treatment of C. jejuni NCTC 11168 biofilms was >80%, whereas bacteriophage resistance was not found in similarly recovered C. jejuni PT14 cells. Concomitant dispersal of the biofilm matrix by bacteriophage was demonstrated by crystal violet staining and transmission electron microscopy (TEM). The resistant survivors of bacteriophage treatment of biofilms formed by HPC5 and PT14 remained closely associated with the phage but not NCTC 11168. Analysis of the DNA contents of these isolates by PFGE and Southern transfer confirmed the presence of phage genomic DNA (approximately 140 kb) leading to the conclusion that these strains represent examples of the carrier state life cycle (CSLC) reported for other bacterial species. TEMs of CSLC cultures demonstrated the association of bacteriophage particles with Campylobacter cells that were devoid of flagella. Physiological studies of the CSLC strains showed the bacteria were non-motile but able to grow at a similar rate to parental cultures until reaching the phage proliferation threshold (7 Log10 CFU/ml) when growth rate declined and the phage titre increased. Of further note the CSLC strains had a greater capacity to survive atmospheric oxygen under nutrient limited conditions. CSLC phages exhibited differences in host binding, efficiency of plating and host range. Transcriptome analyses of CSLC strains harvested from microaerobic cultures at early exponential phase prior to phage proliferation were performed using DNA microarrays to demonstrate changes in host gene expression as compared with parental cultures. Notably genes involved in metabolism and the modification of macromolecules were up-regulated and specific flagella biosynthesis functions down-regulated in the CSLC strains

Bacteriophage treatment of Campylobacter biofilms : formation of the carrier state life cycle

Campylobacter jejuni is a gram-negative thermotolerant microaerobic pathogen that causes human gastroenteritis worldwide. The production of extracellular polymeric substances to create a biofilm is a mechanism by which C. jejuni can protect itself from unfavourable environments, and is a contributory factor to the survival and transmission of the organism to farms animals and into the human food chain. Bacteriophages are natural predators of bacteria that have the potential for use as targeted biocontrol agents with the advantage that they can penetrate and affect bacteria embedded in biofilms. The effects of virulent Campylobacter-specific bacteriophages CP8 and CP30 on C. jejuni biofilms formed on glass by strains NCTC 11168, PT14 and HPC5 at 37 °C under microaerobic conditions were investigated. Independent bacteriophage treatment led to 1 to 3 Log10 CFU/cm2 reductions in the viable count 24 h postinfection compared with control levels. In contrast, bacteriophage applied under these conditions effected a reduction of less than 1 Log10 CFU/ml in planktonic cells. Resistance to bacteriophage in bacteria surviving bacteriphage treatment of C. jejuni NCTC 11168 biofilms was >80%, whereas bacteriophage resistance was not found in similarly recovered C. jejuni PT14 cells. Concomitant dispersal of the biofilm matrix by bacteriophage was demonstrated by crystal violet staining and transmission electron microscopy (TEM). The resistant survivors of bacteriophage treatment of biofilms formed by HPC5 and PT14 remained closely associated with the phage but not NCTC 11168. Analysis of the DNA contents of these isolates by PFGE and Southern transfer confirmed the presence of phage genomic DNA (approximately 140 kb) leading to the conclusion that these strains represent examples of the carrier state life cycle (CSLC) reported for other bacterial species. TEMs of CSLC cultures demonstrated the association of bacteriophage particles with Campylobacter cells that were devoid of flagella. Physiological studies of the CSLC strains showed the bacteria were non-motile but able to grow at a similar rate to parental cultures until reaching the phage proliferation threshold (7 Log10 CFU/ml) when growth rate declined and the phage titre increased. Of further note the CSLC strains had a greater capacity to survive atmospheric oxygen under nutrient limited conditions. CSLC phages exhibited differences in host binding, efficiency of plating and host range. Transcriptome analyses of CSLC strains harvested from microaerobic cultures at early exponential phase prior to phage proliferation were performed using DNA microarrays to demonstrate changes in host gene expression as compared with parental cultures. Notably genes involved in metabolism and the modification of macromolecules were up-regulated and specific flagella biosynthesis functions down-regulated in the CSLC strains

Transformation of Laguna Lake from a marine arm of Manila Bay to a freshwater system

Abstract only.Evolution of Laguna Lake during the past 6,000 years is established using paleontologic, sedimentologic, and geochemical profiles of a 10.5-meter long sediment core from the western lobe. Marine molluscan and diatom assemblages reveal a lagoonal environment with depths initially greater than 10 m, 5,700 radiocarbon years ago (yBP), then it consistently shallowed to intertidal depths until 3,700 yBP. Tellina sp., Leptaxinus sp., and Anomalocardia sp. predominated the molluscan fauna. The marine Cyclotella stylorum, Thalassiosira eccentrica, and Thalassiosira lineata, comprise 80% of the diatom fauna. Brackish water condition was short-lived and it was succeeded by freshwater conditions set at the appearance of Vivipara burroughiana 3,000 yBP. The XRF-derived sulfur, strontium, and calcium concentrations and their aluminum-normalized trends used as geochemical proxy records, corroborate the changing salinity levels and the age boundaries set from the molluscs. Lowering of sea level from the mid Holocene high and vertical movements across the West Marikina Valley Fault, which presently bound the western edge of Laguna Lake, led to the emergence of the Muntinlupa-Paranaque stretch, isolating the proto-Laguna Lake from Manila Bay. Pulsed decreases of salinity to freshwater levels at 250-year intervals and the associated emergence of a 30 m-high barrier reveal a large tectonic influence as opposed to small climatic and sea level shifts within this period. In the last few decades, a new phase of environmental shift has been occurring in the lake. Global warming and land subsidence elevated the saltwater backflow into the lake, enough to leave an increasing salinity trend in the sediment record

Typhoon Haiyan Overwash Sediments From Leyte Gulf Coastlines Show Local Spatial Variations With Hybrid Storm and Tsunami Signatures

Marine inundation associated with the 5 to 8 m storm surge of Typhoon Haiyan in 2013 left overwash sediments inland on the coastal plains of the northwestern shores of Leyte Gulf, Philippines. The Haiyan overwash deposit provides a modern sedimentary record of storm surge deposition from a Category 5 landfalling typhoon. We studied overwash sediments at two locations that experienced similar storm surge conditions but represent contrasting sedimentological regimes, namely a siliciclastic coast and a mixed siliciclastic-carbonate coast. The contrasting local geology is significantly reflected in the differences in sediment grain size, composition and sorting at the two sites. The Haiyan overwash sediments are predominantly sand and silt and can be traced up to ~ 1.6 km inland, extending farther beyond the previously reported \u3c 300 m inland limit of sedimentation. Sites with similar geology, topographic relief, and overland flow conditions show significant spatial variability of sediment thickness and inland extent. We infer that other local factors such as small-scale variations in topography and the type of vegetation covermight influence the spatial distribution of overwash sediments. The Haiyan overwash deposits exhibit planar stratification, a coarsening upward sequence, a non-systematic landward fining trend, and a sharp depositional (rarely erosional) basal contact with the underlying substrate. Overall, the Haiyan deposits have sedimentologic and stratigraphic characteristics that show a hybrid signature common to both storm and tsunami deposits

2016 Philippine Climate Change Assessment (PhilCCA): The Physical Science Basis

This initial assessment of the state of climate change science in the Philippines indicates that climate science in the country is still in its infancy. This report identifies many areas that need further examination, such as the influence of large-scale climate drivers (e.g., ENSO, the Madden-Julian Oscillation, the Pacific Decadal Oscillation) on Philippine climate, the effect of sea level rise on saltwater intrusion and storm surges along coastal areas, and local climate impacts of aerosols and land use change, as well as their interaction with the enhanced greenhouse effect

Complete genome sequence of universal bacteriophage host strain Campylobacter jejuni subsp. jejuni PT14

Campylobacter jejuni strain PT14 is a clinical isolate previously used to propagate bacteriophages in the United Kingdom phage typing scheme. The strain has proven useful in the isolation of Campylobacter bacteriophages from several sources, and it functions as a model host in phage therapy experiments with poultry and poultry meat

The bacteriophage carrier state of Campylobacter jejuni features changes in host non-coding RNAs and the acquisition of new host-derived CRISPR spacer sequences

Incorporation of self-derived CRISPR DNA protospacers in Campylobacter jejuni PT14 occurs in the presence of bacteriophages encoding a CRISPR-like Cas4 protein. This phenomenon was evident in carrier state infections where both bacteriophages and host are maintained for seemingly indefinite periods as stable populations following serial passage. Carrier state cultures of C. jejuni PT14 have greater aerotolerance in nutrient limited conditions, and may have arisen as an evolutionary response to selective pressures imposed during periods in the extra-intestinal environment. A consequence of this is that bacteriophage and host remain associated and able to survive transition periods where the chances of replicative success are greatly diminished. The majority of the bacteriophage population do not commit to lytic infection, and conversely the bacterial population tolerates low-level bacteriophage replication. We recently examined the effects of Campylobacter bacteriophage/C. jejuni PT14 CRISPR spacer acquisition using deep sequencing strategies of DNA and RNA-Seq to analyze carrier state cultures. This approach identified de novo spacer acquisition in C. jejuni PT14 associated with Class III Campylobacter phages CP8/CP30A but spacer acquisition was oriented toward the capture of host DNA. In the absence of bacteriophage predation the CRISPR spacers in uninfected C. jejuni PT14 cultures remain unchanged. A distinct preference was observed for incorporation of self-derived protospacers into the third spacer position of the C. jejuni PT14 CRISPR array, with the first and second spacers remaining fixed. RNA-Seq also revealed the variation in the synthesis of non-coding RNAs with the potential to bind bacteriophage genes and/or transcript sequences

Host adaption to the bacteriophage carrier state of Campylobacter jejuni

The carrier state of the foodborne pathogen Campylobacter jejuni represents an alternative life cycle whereby virulent bacteriophage can persistent in association with host bacteria without commitment to lysogeny. Host bacteria exhibit significant phenotypic changes that improve their ability to survive extra-intestinal environments but exhibit growth phase dependent impairment in motility. We demonstrate that early-exponential phase cultures become synchronised with respect to the non-motile phenotype, which corresponds with a reduction in their ability adhere and invade intestinal epithelial cells. Comparative transcriptome analyses (RNA-seq) identify changes in gene expression that account for the observed phenotypes: down regulation of stress response genes hrcA, hspR and perR; and down regulation of the major flagellin flaA with the chemotactic response signalling genes cheV, cheA and cheW. These changes present mechanisms by which the host and bacteriophage can remain associated without lysis, and the cultures survive extra-intestinal transit. These data provide a basis for understanding a critical link in the ecology of Campylobacter bacteriophage